Mamai et al. Parasites & Vectors (2016) 9:11
DOI 10.1186/s13071-015-1289-0
RESEARCH
Open Access
Morphological changes in the spiracles of
Anopheles gambiae s.l (Diptera) as a
response to the dry season conditions in
Burkina Faso (West Africa)
Wadaka Mamai1,2,3,7*, Karine Mouline1,2, Jean-Philippe Parvy4,5, Jo Le Lannic6, Kounbobr Roch Dabiré1,
Georges Anicet Ouédraogo3, David Renault6 and Frederic Simard2
Abstract
Background: Survival to dry season conditions of sub-Saharan savannahs is a major challenge for insects inhabiting
such environments, especially regarding the desiccation threat they are exposed to. While extensive literature about
insect seasonality has revealed morphologic, metabolic and physiological changes in many species, only a few studies
have explored the responses following exposure to the stressful dry season conditions in major malaria vectors. Here,
we explored morphological changes triggered by exposure to dry season conditions in An. gambiae s.l. mosquitoes by
comparing females reared in climatic chambers reflecting environmental conditions found in mosquito habitats during
the rainy and dry seasons in a savannah area of Burkina Faso (West Africa).
Results: Using scanning electron microscopy (SEM) and confocal imaging, we revealed significant changes in
morphological features of the spiracles in females An. gambiae s.l. exposed to contrasted environmental conditions.
Hence, the hairs surrounding the spiracles were thicker in the three species when raised under dry season environmental
conditions. The thicker hairs were in some cases totally obstructing spiracular openings. Specific staining provided
evidence against contamination by external microorganisms such as bacteria and fungi. However, only further analysis
would unequivocally rule out the hypothesis of experimental artifact.
Conclusion: Morphological changes in spiracular features probably help to limit body water loss during desiccating
conditions, therefore contributing to insect survival. Differences between species within the An. gambiae complex
might therefore reflect different survival strategies used by these species to overcome the detrimental dry season
conditions in the wild.
Keywords: Spiracle, Morphology, SEM, Desiccation, Anopheles gambiae, Burkina Faso
Background
Habitats of tropical savannahs are characterized by
pronounced seasonal and daily fluctuations in environmental conditions such as temperature (with hot days
and cool nights) and relative humidity. The conditions
* Correspondence: [email protected]
1
Institut de Recherche en Sciences de la Santé (IRSS), Direction Régionale de
l’Ouest (DRO), 399 Avenue de la Liberté, 01 BP 545 Bobo-Dioulasso, Burkina
Faso
2
MIVEGEC, UMR IRD 224-CNRS 5290-Université de Montpellier, Institut de
Recherche pour le Développement, 911 Avenue Agropolis, BP 64501, 34394
Montpellier, cedex 5, France
Full list of author information is available at the end of the article
for malaria transmission in these regions are suitable
only during certain periods of the year, particularly in
the rainy season [1]. Thus, the vector dynamics, reproductive period and disease transmission intensity fluctuate
considerably with this seasonal variation in temperature,
precipitation and day length [2–4]. During the unfavourable (dry) weather, malaria mosquitoes of the Anopheles
gambiae s.l. species complex are exposed to severe desiccation challenge, either through the drying-up of their
breeding sites and/or the low ambient relative humidity
[5–7]. Additionally, in early dry season conditions, mosquito larvae might further experience increased crowding
© 2016 Mamai et al. Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
Mamai et al. Parasites & Vectors (2016) 9:11
while available surface water collections shrink and vanish.
To survive through unfavourable conditions, many insects
undergo dormancy (diapause or quiescence), characterized by a suite of morphological, physiological, biochemical and behavioural changes that enhance tolerance and
extend survival to environmental stresses [8, 9] and in particular to desiccation [10].
The two molecular forms of An. gambiae s.s., recently
named An. coluzzii (former M molecular form) and An.
gambiae (former S molecular form) [11] and An. arabiensis are members of the Anopheles gambiae s.l. complex, a group of closely related and morphologically
indistinguishable species [12]. Their distribution ranges
encompass broad environmental and ecological settings,
including arid and semi-arid areas. Although widely
sympatric, the three species exhibit molecular, behavioural, physiological, and ecological differences [13–18].
Despite increased attention drawn in recent years on dry
season survival strategies in these major African malaria
vectors species [7, 19–21], little is known about the processes that sustains survival during the stressful dry season conditions. A recent field study showed evidence
that aestivation (summer diapause) is one mechanism
that allows An. coluzzii to persist in the Sahel [7]. However, migration to/from more favourable localities where
reproduction continues year-round might also be involved [19].
Suppression of water loss is a characteristic of species
that face weather-induced desiccation [22]. It is also
known that the main route of water loss is the cuticle,
while water loss during respiration accounts for about
5–20 % of the total water evaporation [23, 24]. Respiratory gas exchanges in mosquitoes occur through a
multi-branched tracheal system, where cuticular openings called “spiracles” are located on the thorax and the
abdomen. Spiracles are very variable structurally between genus and species, however, typically, the opening
leads to a cavity (the atrium) from which the tracheae
arise. In addition, spiracles of most insects have closing
valves and can be surrounded by dust-catching hairs. In
adult mosquitoes, spiracles are paired, bilaterally symmetric and located on the mesothorax, metathorax, and
abdominal segments. Their apertures ensure the tradeoff between gas exchanges and water loss [7], since oxygen, a necessary gas for cell activity, must pass through
the spiracles to enter the respiratory system [25]. Regulation of spiracle aperture plays a role in water conservation and may best be illustrated in insects showing
discontinuous respiration [26, 27]. Although the adaptive
significance of discontinuous gas exchange (DGC) is a
subject of considerable debate, this respiratory regimen
is characterized by a period in which the spiracles are
fully closed. Indeed, DGC is a repeating cycle of spiracular openings and closings that leads to periodic releases
Page 2 of 9
of carbon dioxide [28]. In ants Pogonomyrmex barbatus,
the metabolic rates were found lowest for individuals
using DGC, intermediate for individuals using cyclic gas
exchange, and highest for individuals using continuous
gas exchange [29]. Permanently opened spiracles allow
maximum gas exchange but insects face desiccation
stress more quickly [30]. Studying the effects of a xeric
environment on water balance in Glossina sp., Bushel
[31] concluded that increased water retention in Glossina sp from xeric environments resulted largely from
spiracular control of transpiration. Spiracle size could
also be positively correlated with water loss. Indeed,
Nagpal and collaborators [32] showed that the spiracles
of ecological variants of An. stephensi displayed different
sizes, being the smallest in the xerophilic ecotype.
Water conservation mechanisms may be of considerable importance to survival and there is evidence that
spiracles are instrumental in water conservation while
still responding to the often-conflicting demands of
respiration. The object of our study was therefore to survey, using electronic and confocal microscopy, the effect
of contrasted environmental conditions on spiracles
morphology of adult females of the An. gambiae s.l.
complex raised under environmental conditions mimicking those found in a savannah area of Burkina Faso during the rainy and the dry seasons.
Methods
Mosquitoes
We used mosquito colonies maintained at the Institut de
Recherche en Sciences de la Santé (IRSS) insectaries in
Bobo-Dioulasso under controlled conditions (27 ± 1 °C,
80 ± 10 % relative humidity (RH) and 12:12 dark/light
cycle). The An. arabiensis colony originated from wild
gravid females collected in Bobo-Dioulasso (11°10’ N, 4°
17’ W) in 2008, the An. coluzzii colony was seeded from
females collected in the village of Bama, Vallée du Kou
(11°23’ N, 4°24’ W) in 2008, and the An. gambiae colony
originated from females collected in the village of
Soumousso (11°04’ N, 4°03’ W) in 2009 (see [18] for further details). All these sites are located in southwestern
Burkina Faso, within 50 km from each other and were
previously described [15, 18, 33].
Environmental conditions, mosquito rearing and sample
collection
Mosquitoes were reared from the egg to the adult stage
in programmable climatic chambers (Sanyo MLR 315H,
Japan), where the climatic parameters characterizing the
rainy and the dry season conditions were defined from
hourly averaged records collected in Bama during
August 2010 (rainy season, RS) (Additional file 1: Figure
S1A) and December 2010 (dry season, DS) (Additional
file 1: Figure S1C) using a Vantage Pro2 monitoring
Mamai et al. Parasites & Vectors (2016) 9:11
station (Weatherlink; Davis Instruments, Hayward, CA,
U.S.A.) As previously described [18, 34], twelve steps cycles were designed to reproduce as close as possible the
natural climatic variations monitored in the fields. A
photoperiod of 12 L: 12D was set, corresponding to day
length at the end of the rainy season in Bama (i.e., mid/
late October), when dry season survival strategies might
be set up in sensitive stages. To control for potential
cycle failure, temperature and relative humidity were recorded inside the chamber at a 10 min pace using
MSR145 Data Loggers (MSR145B4HL, MSR electronics
GmBH, Switzerland). We also recorded the temperature
cycles to which larvae were exposed using waterproof
MSR145B4T2L monitors. The daily air temperature and
humidity fluctuations recorded in the climatic chamber,
identical to [18, 34], are represented in Additional file 1:
Figure S1B and D and water temperature variation is
shown in Additional file 2: Figure S2.
For each mosquito colony, eggs were collected from
three independent batches, each produced by >50 caged
females. Eggs were transferred into transparent plastic
trays (21.5 × 16 × 9.5 cm) containing spring water and incubated in the climatic chambers. After hatching, firstinstars were counted and distributed into new plastic
trays containing 1 L of spring water at a density of 100
larvae per tray. Each species was reared in separate pans,
and three trays were used per colony and per environmental condition. The first two larval instars were fed
every other day with 0.30 mg of ground fish food (Tetramin®), whereas later instars were supplied with 0.75 mg
of food. Water was renewed when necessary to avoid
scum formation and fouling of the media. The water
used for renewal was stored in advance in the climatic
chambers to avoid any perturbation of the temperature
cycles. Every day, the position of the trays was randomly
alternated to avoid positional effects within the incubators. Before adult emergence, pupae were collected with
pipettes and transferred into plastic cups covered with
netting. Emerging adults were immediately removed
with an aspirator. Males were discarded and only females were kept in plastic cups closed with nets and fed
using a cotton ball soaked with 10 % glucose solution.
Starting from the first day after their emergence, the dry
season (DS) females (inside the cups) were placed into
large plastic boxes filled with desiccant (Silica gel
Chameleon©), where relative humidity reaches values as
low as 15 %, as measured by a T/H recorder MSR145
(MSR Electronics, GmBH, Switzerland). This was performed to mimic the field conditions, where relative humidity drops below levels allowed in the climatic
chambers during the hottest hours in December
(Additional file 1: Figure S1A). Hence, females were
placed into the boxes (without access to sugar) at 10 a.m.
and removed at 4 p.m., where a new cotton ball soaked
Page 3 of 9
with glucose solution was provided. For consistency, the
same protocol was applied to the females raised under
rainy season (RS) conditions, except that the boxes were
filled with desiccant soaked with water. Environmental
conditions monitored inside the boxes are provided in
Additional file 1: Figure S1C and D. This was reproduced
each day until the 10th day post emergence, where females of each species were collected around 2 p.m. at the
very same time for each environmental condition to avoid
potential confounding effect of circadian rhythms on spiracle aperture. Samples were snap frozen and preserved in
70 % ethanol under ambient temperature. Twenty
alcohol-preserved specimens were sent to the University
of Rennes 1, France, before being prepared for further
processing through scanning electron microscopy (below).
A second experiment involving An. coluzzii and An.
gambiae raised under the same dry season conditions as
described above was further performed for confocal imaging (see below). This experiment involved 50 ethanol
preserved mosquitoes from each species, which were
sent to the University of Pierre et Marie Curie, Paris,
France for spiracle staining and observations using confocal microscopy.
Scanning Electron Microscopy (SEM)
Eight females from the first experiment were randomly
picked for each species and each environmental condition and processed for SEM observations. The females
were dehydrated by immersion during 30 min in ethanol/water solutions of graded ethanol concentrations
(70, 80, 90, 95 and 100 %), according to [35 ]. Mosquitoes were then incubated in acetone and kept until use.
They were dried by the critical point method using liquid CO2 in a Balzers Critical Point Drier (Balzers
Union FL-9496 Balzers/Furstentum Liechtenstein,
Germany) apparatus attached to specimen holders and
coated with gold and palladium in a sputter coater
(FINE COAT ion sputter JFC-1100, JEOL, Japon).
Finally, the specimens were observed under a scanning
electron microscope (JEOL SJS-6301F, Japan).
Confocal imaging and spiracle staining
Females of the second experiment and those from the field
(for which the species status was assessed using PCR technique [36]) were dedicated to the confocal imaging only,
for which mosquito thoraces were dissected in Phosphate
Buffer Saline (PBS) and fixed for 30 min in PBT (PBS +
0.1 % Triton X-100) containing 4 % of paraformaldehyde.
The thoraces of 25 randomly picked females from experiment 2 (seven from the species An. coluzzii, 18 from
An. gambiae, see Table 1) were further stained with a
mixture containing Phalloidin (a commonly used stain
for F-actin filaments) and TO-PRO®-3 (a commonly used
stain for nucleic acids): after three washes of 10 min in
Mamai et al. Parasites & Vectors (2016) 9:11
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Table 1 Number of opened and closed spiracles and number of spiracles displaying coated setae when observed under confocal
microscope after staining with the vital stain TO-PRO-3. Unobservable spiracles are those for which the dissection and/or mounting
steps before observation were unsuccessful. Observed mosquitoes were from experiment 2
PBT, thoraces were stained overnight at 4 °C in PBT
containing Alexa Fluor® 568 Phalloidin (0.07 μM) and
TO-PRO®-3 (0.01 mM). Samples were rinsed 3 times for
10 min in PBT then hemi-thoraces were mounted with
spiracles facing the coverslip in DABCO (Sigma) and examined by confocal microscopy using a Nikon (TE
2000-U) microscope.
Results
Morphological variation between species and environmental
conditions
Entire females from experiment 1 were carefully observed under the SEM in order to look for striking morphological and structural differences between species
and/or environmental conditions, with a focus on the respiratory system. The only differences that jumped out
during our observations resided in the structural appearance of the spiracle apparatus.
The three species displayed differences in the visual
aspect of the trichomes (or setae) of the mesothoracic
spiracles in females reared in DS conditions compared
to those reared in RS conditions (Fig. 1). Indeed, our observations showed that in 100 % of the females observed
under the SEM, the mesothoracic spiracles are wide
open in females reared in RS conditions whereas the
hairs appear wider and thicker in females reared under
DS conditions, ultimately plugging entirely the spiracular
aperture in An. coluzzii and An. arabiensis (Figs. 1 and 2).
The phenotype is less striking in An. gambiae females,
where the hairs, although oversized, leave a wide aperture.
We were able to observe abdominal spiracles in only 4–5
females per species and conditions; however, all the spiracle structures observed in DS females were modified, and
displayed obstructed apertures (Fig. 1). This suggests that
the morphological modification of spiracular associated
structures applies to the whole respiratory system. We
confirmed our results in a second experiment where
meso- and metathoracic spiracles of 25 females raised
under DS conditions were observed under confocal imaging (Fig. 2, Table 1). Overall, about 63 % of An. gambiae
(N = 18) and 28 % An. coluzzii (N = 7) females displayed
oversized hairs around their thoracic spiracles under dry
season conditions (Table 1). Thickened hairs were not always associated with the mechanical closure of the spiracular valves and both phenotypes can be independent
although a significant trend appears. Of all the spiracle
structures showing oversized hairs, 57 %
(17 out of 30)
were mechanically closed whereas this percentage was
28 % (25 out 90) for the spiracles without coating (χ21 =
7.033, p = 0.008).
Spiracle staining, observations, and related hypothesis on
the origin of the oversized hairs
In all the mosquitoes we observed, neither Phalloidin
(which stains F-actin filaments) nor TO-PRO®-3 (which
stains nucleic acids) probes gave specific signals around
or in the thickened hairs, whereas for TO-PRO®-3, a specific staining was detected in the nuclei of mosquitoes’
muscles (Fig. 3, asterisks). This staining is taken as our
positive control and rules out a potential failure in the
Mamai et al. Parasites & Vectors (2016) 9:11
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Fig. 1 Observation using SEM imaging of mesothoracic and abdominal spiracles of An. gambiae s.l. females raised under different environmental
conditions (DS: Dry Season; RS: Rainy Season). Spiracle features surrounding spiracles are thickened under dry season conditions. White arrows
point to the thickening of the spiracular features. a.: An. arabiensis; (b).: An. gambiae; (c).: An. coluzzii
Fig. 2 Observation using confocal imaging showing a meta-thoracic spiracle with thickened hairs in two females An. coluzzii raised under dry
season conditions
Mamai et al. Parasites & Vectors (2016) 9:11
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Fig. 3 Phalloidin (a, c and e) and TO-PRO-3 (b, d and f) staining of spiracles and thick setae in An. coluzzii females raised under dry season conditions. Spiracle structures were observed using confocal imaging
staining or detection procedures to explain the lack of
signal in the setae.
Over 50 photographs under confocal imaging were
taken that allowed the following observations: (i) in spiracles for which setae are not oversized, we can see that
these structures are rooted in the insect’s cuticle
(Fig. 3e–f ), (ii) setae are present at similar densities in all
spiracles structures, whether they are oversized or not
(Fig. 3f ).
The lack of live cell specific staining, the observation
that the oversized setae are differentiated from the mosquito cuticle and the fact that setae’s densities between
and specimen are equivalent among all the observed
specimens, are all evidence against the hypothesis of this
structure being built by the aggregation of contaminating microorganisms like bacteria or fungi. However, only
further analysis of the qualitative nature of these structures would unequivocally rule out the hypothesis of experimental artifact.
Discussion
Despite the epidemiological importance of Anopheles
mosquitoes, little is known about the mechanisms
underpinning the survival of these species during the
harsh conditions of the dry season in Africa. Here, we
provide evidence for morphological variation in species
of the An. gambiae s.l. complex in response to environmental variations. Because desiccation threat is more
Mamai et al. Parasites & Vectors (2016) 9:11
severe when spiracles are open [37, 38], we expected
that mosquitoes reared in low ambient humidity and
high temperatures, as typically observed during the dry
season in sub-Saharan Africa, would exhibit morphological traits that serve as adaptations to reduce water
loss. In fact, water is lost rapidly through opened spiracles
during respiration [12, 39, 40]. In the arid-adapted ant
Cataglyphis bicolor, the thoracic spiracles act as highcapacity gateways to the tracheal system and are responsible for approximately 90 % of the total gas exchange
including water vapour during running activity [41].
Moreover, studies from other insects have demonstrated
that spiracle closure greatly facilitates water conservation
[37, 42–45]. For example, by measuring the rate of water
loss of tsetse flies in varying states of desiccation, Bursell
[37] was able to show that spiracular control of transpiration increased as water reserves decreased.
The ability to close spiracles mechanically using the
valve mechanism is a physiological adaptation that reduces water loss in insects [38, 46–48]. Hence, closed
spiracles have been reported in Drosophila melanogaster
during flight to reduce water loss and gas exchange into
the tracheal system [38]. Although functional experiments are lacking to link the size or shape of the thoracic or abdominal spiracles with heat and/or desiccation
tolerance, it is nonetheless clear that spiracular structures evolve towards smaller sizes in mosquito species
inhabiting dry climates [49]. For instance, in the hygrophilous species of Anopheles and Aedes, spiracles are
generally larger, whereas in the xerophilous species the
openings are much smaller [30, 49]. Moreover, the spiracular index, i.e. the ratio of the length of the spiracle to
the length of thorax has been used as a tool to identify
ecological variants exposed to contrasted environments
in An. stephensi, this ratio being smaller in xeric vs
mesic environments [32].
The mechanical closure of spiracles in desiccating conditions represents a rapid and transient response, which
is thought to be regulated by a sensory mechanism elicited by low relative humidity [32]. In contrast, the specific spiracular structure modifications we observed
through the whole respiratory system might represent
important long-term adaptation, which could be programmed in anticipation to DS conditions. Hence, the
ability to build oversized hairs that ultimately plug the spiracle apertures could be triggered by environmental cues
sensed during the aquatic stages (i.e. high temperatures) or
right upon emergence of adult mosquitoes (i.e. high temperatures and/or low relative humidity). Trichomes of various size, shape and density have been reported to line the
spiracles atrium cavity in Geophilomorpha species, these
structures being described as “solid and expended distally
and showing a network of sclerotisation” in Cormocephalus calcaratus, “flap-like” in Strigamia, “cone shaped” in
Page 7 of 9
Geophilus insculptus, or “elongated plates” in Hapiophilus
subterraneous (Lewis, [50] and references therein). In his
attempts to provide a functional explanation to such variability, Lewis [22], reported a strong correlation between
the presence of a thick layer of trichomes in the atrium
and resistance to desiccation challenge in 4 geophilomorphs. He concluded that this structure, together with a
narrowed spiracular opening, might limit water evaporation from the atrium. Outgrowths cover the spiracular
openings of the xerophilous buprestid beetle and, in the
cockroach, such structures line the atrium, either outside
or inside the valves (Hadley, [23] and references therein).
Oversized setae around the spiracles may therefore well
play the same role in mosquitoes than in the above-cited
species, and might contribute to maintaining minimal
metabolic activity while minimizing respiratory water loss.
As such, they could be part of the survival strategies developed by An. gambiae s.l. to cope with low relative humidity
values encountered during the dry season, either under
diapause, quiescence or reproductively active states.
However, to our knowledge, our study is the first to report trichomes variability within the same species, putatively induced by environmental triggers. Dry season
specific phenotype of the spiracles’ setae was also observed in preliminary experiments involving mosquitoes
collected in Bama at the beginning of the dry season.
Two out of 22 mosquitoes observed under confocal imaging were showing oversized setae (i.e. 9 %), however,
the phenotype was less striking in the sense that very
few setae were indeed oversized (data not shown), which
might suggest a progressive building of the phenotype
over time, either with mosquito age and/or with the installation of the dry season. These preliminary observations warrant further investigations under field and/or
semi-field conditions that will help resolve the biological
meaning of such phenotypes as well as its underlying
physiological and ecological determinants. Nonetheless,
given that such structural variation has not been observed in any Anopheles mosquito to date, further experiments must also be conducted to strengthen our
findings and to definitely rule out the caveat of an experimental artifact. Among these, a time course observation of the progressive building of oversized trichomes,
careful observation of a recent study on Drosophila melanogaster revealed that lipid deposits are used to waterproof the spiracles [51], a link that might also exist in
An. gambiae s.l. mosquitoes.
The ability to manage water reserves through the
modification of spiracular morphology might account, at
least in part, for better survival under desiccation challenge and increased body water content observed in
mosquitoes when raised under dry season conditions
[52]. In addition to water conservation, reduced spiracle
openings might also contribute to lower metabolic rate
Mamai et al. Parasites & Vectors (2016) 9:11
and gas exchange through respiration. Huestis and collaborators [20] indicated that the mean metabolic rate of
An. coluzzii was lowest during the transition period between the wet and the dry season in the Sahel, which is
consistent with obstructed spiracles as we observed in
mosquitoes reared under DS conditions.
Conclusion
This study identified morphological variations in An.
gambiae s.l. mosquitoes when exposed to the severe dry
season conditions in West African savannahs. These
morphological changes might reflect specific adaptations
to increase survival under different climatic or microclimatic conditions, pointing towards an important influence of spiracles’s hairs on the rate of respiratory water
loss and slowing down of the global metabolism. Although
there is clear evidence for seasonal differentiation in An.
gambiae s.l. species, further research including cuticle
morphology and composition and gas exchange rates are
required to explore in more detail the biological relevance
and adaptive value of these morphological adaptations.
Additional files
Additional file 1: Figure S1. Daily environmental conditions of
temperature (dashed line), relative humidity (solid line), and dark/light
duration (horizontal bars) recorded during the wet season in the field (A)
inside the climatic chambers (B) and at the onset of the dry season in
the field (C) and inside the climatic chambers (D). The red arrows indicate
the times when females inside the cups were placed into large plastic
boxes filled with desiccant (10 h) and were removed (18 h). Between the
arrows, temperature and relative humidity values are those recorded
inside the plastic boxes. Humidity values inside the climatic chamber are
2–5 % above field ones for the rainy season conditions due to condensation
problems in climatic chambers at higher humidity values. (TIF 870 kb)
Additional file 2: Figure S2. Daily temperature of the larval rearing
water inside climatic chambers; wet season conditions (dashed line) and
dry season conditions (solid line). (TIF 547 kb)
Abbreviations
DS: Dry Season; RS: Rainy Season; IRSS: Institut de Recherche en Sciences de
la Santé; WHO: World Health Organisation; RH: Relative Humidity;
SEM: Scanning Electron Microscopy.
Competing interests
The authors have declared that no competing interests exist.
Authors’ contributions
KM, DR and FS, conceived the study and coordinated its implementation.
RKD, GAO and JPP participated in the design. WM, KM and JPP performed
the experiments and drafted the manuscript which was critically revised by
DR, FS and RKD. All authors read and approved the final version of the
manuscript.
Acknowledgments
This work is part of the project “Dry Season Survival Strategies in major
African malaria vectors” funded by the French Agence Nationale de la
Recherche through grant N° ANR-08-MIEN-006 to F.S. WM is supported by a
PhD fellowship from the IRD/DSF through “Bourse de soutien de thèse”
program.
Page 8 of 9
Author details
1
Institut de Recherche en Sciences de la Santé (IRSS), Direction Régionale de
l’Ouest (DRO), 399 Avenue de la Liberté, 01 BP 545 Bobo-Dioulasso, Burkina
Faso. 2MIVEGEC, UMR IRD 224-CNRS 5290-Université de Montpellier, Institut
de Recherche pour le Développement, 911 Avenue Agropolis, BP 64501,
34394 Montpellier, cedex 5, France. 3Université Polytechnique de
Bobo-Dioulasso (UPB), Bobo-Dioulasso, Burkina Faso. 4Université Pierre et
Marie Curie, 75005 Paris, France. 5CGM, UPR 3404, CNRS, 91190 Gif-sur-Yvette,
France. 6Université de Rennes 1, UMR CNRS 6553 ECOBIO, Campus de
Beaulieu, 263 Avenue du Gal Leclerc, CS 74205 35042 Rennes, Cedex, France.
7
Institut de Recherche pour le Développement, Antenne de Bobo Dioulasso,
BP 171 Bobo Dioulasso 01, Burkina Faso.
Received: 1 December 2015 Accepted: 25 December 2015
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